Several molecular mechanisms seem to be involved in agonist-mediated receptor desensitization. The cytoplasmic COOH-terminus of the receptor contains a large number of serine and threonine residues, which could be potential phosphorylation sites by protein-kinase C (PKC) or Beta- adrenergic receptor kinase (betaARK). The phosphorylation of thrombin receptor is very likely to occur, but has not been directly proved yet, nor is it clear how phosphorylation is connected with desensitization of the receptor. Moreover it would be interesting to test if phosphorylation of thrombin receptor may play a role in its desensitization and internalization. Nevertheless, the exact residue(s) of phosphorylation is yet to be identified. The aim of this study is to test these possibilities, and to clarify the role of phosphorylation/dephosphorylation in the working model of the thrombin receptor. A direct evidence of phosphorylation of the thrombin receptor in the whole cell would be provided by labeling cells with 32P and by immunoprecipitation of the thrombin receptor. Furthermore, the identification of essential serine residue(s) undergoing phosphorylation could be achieved by using mutant receptors lacking potential sites of phosphorylation and by testing the possibility to be phosphorylated following thrombin-induced activation of cells expressing them. This approach would also allow to clarify the role played by phosphorylation in the thrombin receptor regulation by testing the possibility of desensitization and desensitization of mutant receptors by both thrombin and synthetic peptides. An additional aim of our study is to establish which kinases(s) might be involved in this phenomenon. PKC phosphorylates and desensitizes other G protein-coupled receptors. PKC activated by phorbol esters prevents HEL cells from responding to thrombin and to TRP12/55, but also to neuropeptide Y. Moreover, PKC inhibition by neurosporine only slightly prevents the desensitization by thrombin and TRP12/55, suggesting that PKC might have a limited, if any, role in the desensitization of thrombin receptor.